Necrotic cell death underlies the pathology of many neurodegenerative diseases and is an incapacitating outcome of stroke, ischemia and physical injury. Relatively little is known about genetic control of necrosis, although such mechanistic understanding is clearly critical for effective intervention. We are pursuing a genetic dissection of necrosis mechanisms in the facile model system C. elegans. We have found that hyper-activated ion channels that conduct excess Na +or Ca +2 into neurons initiate a necrotic-like cell death in nematodes (similar events initiate human necrosis). Our analysis of necrosis suppressor loci has identified the ER Ca +2storing protein calreticulin as essential for progression through necrosis. Likewise, the activity of ER Ca +2 release channels potentiates necrosis. We suggest that a rise in intracellular calcium, dependent upon ER Ca +2 stores, is critical for neurotoxicity. We have also identified specific Ca+2-activated calpain proteases and cathepsin proteases as key contributors to necrosis. These may act in a proteolytic pathway to execute cell death. Importantly, intracellular Ca +2 rise and activation of calpain proteases are features of human necrosis, suggesting necrotic death mechanisms are conserved from nematodes to humans. Our Iong term goals are to identify genes that facilitate progression through necrosis and to expand/test a working model in which excess Na + influx signals ER Ca+2 release, with the consequent intracellular Ca +2 rise activating a proteolytic cascade involving specific calpains and cathepsin proteases. We will: 1) use RNAi technology to identify all C. elegans genes needed for efficient progression through necrosis; 2) couple in vivo reporting of intracellular Ca +2 with genetic manipulations to better characterize changes in intracellular Ca+2 that accompany necrosis and to define the critical molecular origins of these changes; 3) use genetic and cell biological approaches to describe mechanistic details of the proteolytic component of necrosis. Overall we expect to exploit unique features of C. elegans to provide a detailed mechanistic description of necrosis, including identification of novel genetic factors influencing this process that may suggest new strategies for limiting devastating effects of necrosis in human injury and disease.